There are two major objectives of this grant. The first is to provide a comprehensive training experience for the principal investigator in the areas of molecular biology and bacterial pathogenicity. The second objective is to apply this expertise to the genetic analysis of surface antigens of the obligate intracellular parasitic bacterium Rickettsia prowazekii. During Phase I the candidate will complete a series of formal courses in molecular and cell biology, genetics, and bacterial pathogenicity. In addition, laboratory rotations will expose the candidate to a variety of research problems and techniques. This particular part of Phase I will culminate in a research rotation at the Centers for Disease Control. Phase I research will be directed toward the characterization of a rickettsial DNA fragment previously shown to express a rickettsial antigen in Escherichia coli. The antigen gene will be localized, subcloned into appropriate vector systems and sequenced. The regulatory regions will be identified and their activity examined. This information in conjunction with direct analysis of the protein will be used to examine the location and function of this antigen. In the last quarter of Phase I the candidate will initiate the production of antigen and the preparation of antiserum for use in Phase II. During Phase II the candidate will initiate the isolation and characterization of rickettsial DNA fragments coding for surface antigens of R. prowazekii. Using antiserum raised against outer membranes of R. prowazekii, previously established cosmid and plasmid clone banks will be screened for the expression of rickettsial antigens. The candidate will also establish phage vector (lambda gt11) banks to help ensure expression of rickettsial epitopes. Clone banks also will be screened with human convalescent serum to identify those antigens to which the human responds during the disease process. Fragments expressing rickettsial surface antigens, with an emphasis on those expressing antigens recognized by the human sera, will be sequenced in order to deduce the protein sequences and to identify the regulatory elements controlling the expression of the antigens. Finally, complement genes and gene products from an avirulent and a virulent strain will be compared in order to identify any potential virulence- associated differences.